The cloning and engineering of genetically encoded fluorescent proteins has allowed the development of many unique tools for studies in cell biology, medicine, and physiology. When combined with intravital microscopy, these probes have the potential to report events from inside living cells in the intact organism. However, the utility of the fluorescent protein biosensors for intravital microscopy has been limited by the poorly characterized behaviors of fluorescent proteins under two-photon excitation. The goal of the Multiphoton Biosensor Development core is to develop and rigorously validate a set of biosensor probes optimized for intravital multiphoton microscopy studies of renal function. Modular probes developed by the core will be incorporated into a toolkit of probes and protocols that will be provided to renal investigators. These probes will provide renal researchers with powerful new tools that will enable them to conduct novel intravital microscopy studies addressing fundamental issues of renal physiology, cell biology and pathophysiology.